DESCRIPTION (Adapted from applicant's Description): The study of calcium signaling in single cells is dramatically increasing our knowledge of cellular physiology. A wealth of information has been gathered on the mechanisms underlying the calcium signal and its role in cellular signal transduction. Recent technological innovations in single-cell measurements make it possible to expand this knowledge base by direct testing of many fundamental hypotheses of cell signaling. Microanalytical chemical separations enable a spatially and temporally defined quantitative measurement of key enzymes and second messengers in signal networks. Imaging of GFP-tagged proteins provides real-time information on changes in localization of many of these molecules. The present work will combine these technologies with the monitoring of intracellular calcium concentrations for in vivo studies of the activation of calcium-dependent enzymes, and of the changes of inositol 1,4,5-trisphosphate during the Xenopus laevis fertilization calcium wave. The specific aims are 1) to measure the spatial and temporal relationship of the activation of calcium-sensing enzymes in vivo to the fertilization calcium wave, and 2) to identify the mechanisms underlying the propagation of the fertilization calcium wave.